| Objective:The chitosan coating was fixed on the surface of the new type of implant "Tixos".The coating's influence on the Schwann's early adherency,proliferation and secretion was tested by the experiment of in vitro cell.Method:Chitosan was fixed on the surface of “Tixos” by cross-linking agent to produce the coating.The experiment was divided into three groups: the cell group used as blank control group,Tixos-CS group used as negative control group and Tixos-CS group used as experimental group.Material's super facial instracture of experimental group and negative control group was observed by SEM.Banding angle of both groups was tested to confirm its hydrophilia.The binding intensity between the chitosan coating and the surface of “Tixos” was tested by the tensile testing machine,experimental materials were soaked in vitro and weigthed every day to measure the degradation time.Materials and Schwann cells were caltured for 12 hours,the amount of cells adherened on the surface of materials was tested by the method of DAPI.Cell morphous on the surface of material was observed by SEM after being cultured for 24 hours.Material's influence on the schwann's proliferation was tested on the first,third and fifth day by CCK-8.After cells and materials' being cultured together,the number of cells was counted.The cell's growing curve was drawn and cell's growing state was observed.Cell cycle tested by Flow cytometry.The condition of schwann's secreting NGF and GPNF was tested by ELISA on the first,second and third day.Statistical analysis was made by SPSS 24.0.Result:1.Observed by SEM,the size of a chitosan particle is about 2-5um,rectangle or polygon.The particles arranged closely and adhered to the surface of “Tixos” tightly.2.The touching angle of the surface of Tixos-CS material is smaller than 90°and the surface shows hydrophilia.3.Tested by the tensile testing machine,chitosan coating could adhere to the surface of “Tixos” steadily.4.The experiment of degradation shows the coating can degrade slowly and uniformly and the degradation was finished about on the eighth day.5.The schwann cells and two groups of materials were cultured together for 12 hours and tested by DAPI,the result showed there were more cells to adhere in the group of Tixos-CS.6.After being cultured for 24 hours,cell's morphous could be observed by SEM.It was seen that pseudopodium stretching out on the surface of materials were more and longer cells stretched more fully.7.As a consequence of CCK-8 test,the absorbance value of Tixos-CS is always higher than that of common Tixos group on the first,third and fifth day.8.Seen from the growing curve,in the group of Tixos,from the first day to second day after cell's passage,it is the period of growing slowly.From the third day to the sixth day,it's the period of growing fast.After the sixth day,it is the flat period.In the group of Tixos-CS,the first day,is the period of growing slowly.From the second day to the fifth day,it is the period of growing fast.After the fifth day,it is the flat period.9.Cells cyde was analysed by flow cytometry: schwann percentade in the period of S+G2/M of Tixos-CS increased compared with that of Tixos on the first and third day.10.It is showed that by the result of ELISA test NGF and GDNF secreted by schwann cells on the Tixos-CS were higher than those of Tixos.Conclusion:1.Tixos-CS can contribute to adherency and proliferation of schwann cells.2.Tixos-CS can contribute to schwann cell's secreting NGF and GDNF. |
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